Recent evidence suggests that a central mechanism may be contributing to the sympathetic abnormality in diabetes. Nitric oxide (NO) has been known as a neurotransmitter in the central nervous system. The goal of this study was to examine the role of the endogenous NO system of the paraventricular nucleus (PVN) in regulation of renal sympathetic nerve activity (RSNA) in streptozotocin (STZ)-induced diabetic rats. The change in number of NADPH-diaphorase positive neurons (a marker for neuronal NO synthase (nNOS) activity) in the PVN was measured. Diabetic rats were found to have significantly fewer nNOS positive cells in the PVN than in the control group (120±11 vs 149±13, P<0.05). Using RT-PCR, western blotting and immunofluorescent staining, it was also found that nNOS mRNA expression and protein level in the PVN were significantly decreased in the diabetic rats. Furthermore, using in vivo microdialysis technique, we found that there was a lower NOx release from the PVN perfusates in rats with diabetes compared to the control rats (142±33nM vs 228±29nM, P<0.05). In -chloralose and urethane anesthetized rats, an inhibitor of NO synthase, L-NMMA microinjected into the PVN produced a dose-dependent increase in RSNA (35±3% vs 11±3%, P<0.05), mean arterial pressure (MAP) and heart rate (HR)in both control and diabetic rats. These responses were significantly attenuated in rats with diabetes compared to control rats. On the other hand, an NO donor, sodium nitroprusside (SNP), microinjected into the PVN produced a dose-dependent decrease in RSNA, MAP and HR in the control and diabetic rats. RSNA (17±3%, vs 41±6%, P<0.05) and MAP in response to SNP were significantly blunted in the diabetic group compared to the control group. In conclusion, these data indicate an altered NO mechanism in the PVN of diabetic rats. This altered mechanism may contribute to the increased renal sympathetic neural activity observed in diabetes.